PROJECT SUMMARY Worldwide, about 390 million people are infected annually with any of the four dengue virus (DENV1-4) serotypes, all of which are capable of eliciting disease. DENV is a mosquito-transmitted member of the flavivirus family, and causes disease ranging from fever and aches to hypovolemic shock and death. To reduce the burden of disease, development of a vaccine which induces a balanced tetravalent response to and protects against all four serotypes simultaneously is a current priority. The proposed studies will address two central issues hampering DENV vaccine development: the lack of a validated correlate of protection and the recent finding that a DENV vaccine may only effective in certain DENV-primed subpopulations. The central hypothesis is that an effective DENV vaccine will elicit adaptive immune B and T cells that recognize such pan-serotype epitopes which are key to protection. Since some of these epitopes are conserved across DENV and other flaviviruses, prior flavivirus exposures can possibly serve to augment these responses and contribute to DENV vaccine protection. To address this hypothesis, samples and clinical data will be used from controlled dengue human infection models that employ tetra- or trivalent vaccination with the NIH's live attenuated DENV followed by challenge with an underattenuated DENV. Samples will also be employed from a trial wherein tetravalent DENV vaccination was administered to subjects who were either flavivirus-nave or had been previously exposed to a flavivirus prior to DENV vaccination. Aim 1 will determine the extent to which pan-DENV B cells and their Ab products which potently recognize DENV1-4 correlate with clinical protection (viremia, rash, blood cell counts, symptoms) by dengue vaccination either in conjunction with or as a complement to the traditional measure of immunogenicity, serum neutralizing antibodies. Aim 2 will measure pan-DENV reactive CD8 and CD4 T cells and determine how the magnitude, kinetics, or phenotype of these responses correlate with protection in relationship to neutralizing antibody. Aim 3 will determine how flavivirus priming affects these B and T cell responses after tetravalent DENV vaccination. Collectively, the proposed studies will greatly expand our understanding of dengue immunogenicity and key factors mediating protection. These studies are innovative in their use of controlled infection and vaccination studies and for the unifying theme of cross-protection at the cellular level. These studies will advance dengue vaccine development by providing new immune markers for further evaluation in field efficacy trials. This project synergizes with the current Translational Global Infectious Disease Research COBRE application by using samples available from its Vaccine Testing Center, by being a data engine to generate novel markers by which to evaluate efficacy in the Mathematical and Computational Predictive Modeling Core, and by leveraging the Human Population Research Core to implement this knowledge into new trials for DENV and eventually for Zika virus.